Numerous approaches have been examined for the potential treatment of Alzheimer's Disease (AD). Despite this massive effort, only two classes of drugs have been approved by the FDA for treatment of AD: cholinesterase inhibitors and memantine. Neither treatment targets the early or causative events in AD and neither is highly effective in preventing disease progression. Thus, there is a critical need for new therapies that target earlier stages of AD, and can be used as a prophylactic measure.
The two pathological hallmarks of AD are the accumulation of intracellular neurofibrillary tangles and extracellular amyloid plaques (also known as senile or neuritic plaques). The major constituent of the amyloid plaque is a peptide known as Aβ or β-amyloid1,2, and aggregation of this peptide is now accepted as a causative factor in the disease3. The Aβ peptide arises via cleavage of the amyloid precursor protein (APP), a single transmembrane protein that is a substrate for several membrane bound proteases. Sequential processing by β- and γ-secretase results in production of Aβ. APP can also be cleaved by α-secretase. However, this cleavage occurs within the Aβ peptide sequence and precludes production of the amyloidogenic Aβ peptide. The opposing results of α- and β-secretase action imply that Aβproduction (and AD progression) could be limited by interventions that shift APP processing away from the β-secretase pathway and/or toward α-secretase.
The processing of APP by α-secretase occurs at the cell-surface4,5. In contrast, β-secretase cleavage occurs within cholesterol-rich membrane domains known as lipid-rafts6-9. The distinct localization of the α- and β-secretase suggests that anti-amyloidogenic or α-cleavage of APP could be accentuated by retention of the APP substrate within membranes of the extracellular surface. Indeed, Aβ production is reduced when APP internalization/endocytosis is blocked by antibody-crosslinking6. Similarly, reduction of APP endocytosis by depletion of membrane cholesterol also attenuates Aβ production4. These findings suggest that AD could be treated with drugs that modulate cholesterol levels or cholesterol-trafficking in the brain.
As described above, APP processing requires an initial cleavage by either α- or β-secretase followed by a secondary cleavage with γ-secretase3. While the first cleavage is critical for establishing the amyloidogenic fate of APP, γ-secretase is also critical as it produces the final Aβ peptide. It appears that γ-secretase activity may also be modulated by membrane cholesterol10,11. For example, cholesterol content in late endosomes/lysosomes is regulated by Niemann-Pick C1 protein (NPC1), and mutations in NPC1 leads to accumulation of unesterified cholesterol in late endosomes/lysosomes12,13. Mice expressing the mutated NPC1 show no changes in amounts of α- or β-secretase activity but exhibit increased γ-secretase activity and accumulate Aβ14,15. Similarly, blockade of lysosomal cholesterol trafficking in neurons also increased γ-secretase activity16. Thus, reduction in membrane cholesterol content can attenuate both the β- and γ-proteolytic pathways which act sequentially to generate Aβ.
The role of cholesterol in the pathogenesis of AD and/or vascular dementia came to the forefront in 2000, when two groups independently reported that subjects that were treated with statins (3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitors) had a significantly lower prevalence of AD. In both studies, statins decreased AD outcome by about 70%17,18. Statins act by inhibiting synthesis of mevalonic acid, a rate-limiting step in cholesterol biosynthesis. In support of a specific epidemiologic connection to cholesterol, several studies have reported an association of mild hypercholesterolemia19-22, high LDL22-25, and low HDL levels23,25,26 with risk for vascular dementia and AD.
Hypercholesterolemia and low HDL levels are established risk-factors for atherosclerosis and atherosclerosis is in turn associated with an increase in AD-pathology. For example, subjects who died of atherosclerotic heart disease without any other neurological diseases had a significantly greater number of amyloid plaques compared to age-matched controls without atherosclerosis27. The extent of arterial stenosis due to atherosclerotic plaques in large CNS blood vessels was also associated with AD28,29. This highlights the overlap between AD, vascular dementia and atherosclerosis.
Recent studies have demonstrated that the nuclear receptors LXRα and LXRβ are central players in cholesterol and triglyceride homeostasis30-32. LXRα is abundant in metabolic tissues including liver, intestine and macrophages. LXRβ is ubiquitously expressed. Both receptors bind to and are activated by cholesterol-derived oxysterol ligands. Activation of LXRs promote cholesterol efflux from many cell-types, including primary astrocyte cultures, primary neurons, and brain-derived cell-lines33-36. Enhanced efflux is mediated by LXR-dependent induction of cholesterol/lipid transporters from the ATP-Binding Cassette family of proteins (ABCA1, ABCG1, ABCG5 and ABCG8). These transporters stabilize and transfer cholesterol to acceptor proteins apolipoprotein-A1 (apoA1) and apolipoprotein E (apoE)37-40. Effects of LXR agonists against both LXRα and β are significant: they increase cellular cholesterol efflux; raise plasma HDL cholesterol; stimulate cholesterol excretion into the bile and feces, decrease hepatic cholesterol content and can reduce atherosclerotic lesions by ˜50% in various murine models41-45. While LXR agonists against both LXRα and β appear to have significant therapeutic potential, they are undesirably associated with a clinically limiting side-effect: Treatment of C57BL/6 mice with T0901317, a LXRα/β pan-agonist induces severe hypertriglyceridemia after only 1 week of treatment45. Therefore, there is a need to identify and use LXR ligands which will dissociate these side-effects from the otherwise positive effects on cholesterol efflux.
Although both LXR-subtypes share a large degree of structural and functional similarity, their distinct tissue distribution profiles imply unique biological activities for each subtype. For example, LXRα knockout mice show reduced plasma triglyceride levels as well as reduced hepatic mRNA levels for enzymes of fatty acid synthesis46. No such effect is observed in LXRβ-null mice. Thus, the hypertriglyceridemia appears to be mediated by an LXRα-dependent activation of hepatic target genes involved in fatty acid synthesis. These target genes include Fatty Acid Synthase (FAS) and SREBP 1c47, a master-regulator of fatty acid synthetic enzymes48. Conversely, ABCA1 is still induced in macrophages from LXRβ-null mice49. Taken together, these findings suggest that LXRα is the subtype responsible for the side-effects of hypertriglyceridemia, whereas activation of LXRβ is sufficient for the positive effects on ABCA1 transcription and macrophage cholesterol efflux.
These observations suggest LXRβ-selective agonists will be particularly useful for the modulation of human lipid metabolism. In particular, LXRβ-selective agonists will maintain the therapeutic effects of LXR activation without promoting the side-effects of hypertriglyceridemia. Therefore, there is a need to identify LXRβ-selective agonists and use LXRβ-selective agonists in the treatment of diseases associated with LXR (e.g., lipid metabolism disorders, artherosclerosis, Alzheimer disease, and inflammation30).